Hearing loss is debilitating and permanent, affecting < 66% of the population over age 70 (Cruickshanks et al., Am. J. Epidemiol., 1998), causing interpersonal isolation and having significant financial impact. During the previous funding period, we used positional cloning techniques to identify mutations in Atp2b2 and Spnb4 in the mouse mutant's deafwaddler and quivering, respectively (Street ea, 1998; Parkinson ea, 2001). For each locus we identified multiple alleles with different degrees of functional impairment. For deafwaddler, the dfw allele is a hypomorph with function reduced to 30% (Penheiter ea) while dfw 2J and dfvv 3J are apparent null alleles with frameshift mutations yielding no detectable protein. We have found that homozygous mutants of all alleles are deaf (Konrad-Martin, 2001 ; Mccullough and Tempel, submitted). Further studies have shown that in homozygous null mice (dfw2J/dfw 2J) endolymphatic calcium concentration drops from 23 mu/M in control to 6 mu/M (Wood et al., 2004). This significant drop in endolyphatic calcium may be causally related to profound deafness, to the degeneration and loss of otoconia (Fonseca-Burke et al., in prep.), as well as to hair cell loss (Pujol et al., 2001) in homozygous null deafwaddler alleles. In heterozygotes no hair cell loss is seen yet hearing loss occurs at high frequencies (Konrad-Martin, 2001; McCullough and Tempel, submitted). These data suggest that calcium is tightly regulated in the inner ear and that PMCA2 (the protein product of the Atp2b2 locus) is a critical player in maintaining calcium balance. Here we propose to extend our studies on deafwaddler by examining its contribution to age-related hearing loss (AHL) and noise-induced hearing loss (NIHL) in mice with graded reductions in Atp2b2 expression. To establish a rationale for comparing man with mouse, we will examine the distribution of PMCA2 in human temporal bone and ask whether patients carrying a deletion of the ATP2B2 locus show hearing loss. Toward identifying pharmacological strategies for preventing hearing loss, we will examine the molecular basis of Atp2b2 gene regulation and ask whether increased levels of expression of Atp2b2 can prevent AHL in mice and protect them from NIHL.